LOCUS       Exported                7372 bp DNA     circular SYN 28-SEP-2021
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    pcDNA3-mRFP-PAGFP-RhoA-Q63L
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7372)
  AUTHORS   Subauste MC, Von Herrath M, Benard V, Chamberlain CE, Chuang TH, Chu
            K, Bokoch GM, Hahn KM
  TITLE     Rho family proteins modulate rapid apoptosis induced by cytotoxic T 
            lymphocytes and Fas.
  JOURNAL   J Biol Chem. 2000 Mar 31. 275(13):9725-33.
  PUBMED    10734125
REFERENCE   2  (bases 1 to 7372)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Jun 7, 2022 from SnapGene Viewer 6.0.5
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "J Biol 
            Chem. 2000 Mar 31. 275(13):9725-33."
FEATURES             Location/Qualifiers
     source          1..7372
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     377..396
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(615..719)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(794..813)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        985..1364
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1365..1568
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1519..1539
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     1613..1632
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1613..1631
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     mRNA            1647..2321
                     /label=mRFP
     CDS             2343..3059
                     /codon_start=1
                     /product="photoactivatable GFP variant (Patterson et al., 
                     2002)"
                     /label=PA-GFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFSYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTMVNRIELKGIDFKEDGNILGHKLEYKYNSHNVYIMADKQKNGIK
                     ANFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSHQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2388..2409)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2649..2668)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2996..3017
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             3066..3647
                     /codon_start=1
                     /product="constitutively active mutant of human Rho family 
                     small GTPase RhoA"
                     /label=RhoA (Q63L)
                     /translation="MAAIRKKLVIVGDGACGKTCLLIVFSKDQFPEVYVPTVFENYVAD
                     IEVDGKQVELALWDTAGLEDYDRLRPLSYPDTDVILMCFSIDSPDSLENIPEKWTPEVK
                     HFCPNVPIILVGNKKDLRNDEHTRRELAKMKQEPVKPEEGRDMANRIGAFGYMECSAKT
                     KDGVREVFEMATRAALQARRGKKKSGCLVL"
     promoter        complement(3675..3693)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(3676..3693)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(3713..3730)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3719..3943
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3989..4417
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4076..4095)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4286..4307
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4426..4446)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4431..4760
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4611..4746
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4673..4692
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4827..5621
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4881..4900)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5491..5510
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5795..5916
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5832..5851)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5886..5905
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5965..5981)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5965..5981)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5978..6000)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5989..6005
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(6013..6043)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6058..6079
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6196..6213)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6367..6955)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6447..6466)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7126..7372,1..614))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
ORIGIN
        1 aattgttgcc gggaagctag agtaagtagt tcgccagtta atagtttgcg caacgttgtt
       61 gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc
      121 ggttcccaac gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa agcggttagc
      181 tccttcggtc ctccgatcgt tgtcagaagt aagttggccg cagtgttatc actcatggtt
      241 atggcagcac tgcataattc tcttactgtc atgccatccg taagatgctt ttctgtgact
      301 ggtgagtact caaccaagtc attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc
      361 ccggcgtcaa tacgggataa taccgcgcca catagcagaa ctttaaaagt gctcatcatt
      421 ggaaaacgtt cttcggggcg aaaactctca aggatcttac cgctgttgag atccagttcg
      481 atgtaaccca ctcgtgcacc caactgatct tcagcatctt ttactttcac cagcgtttct
      541 gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa
      601 tgttgaatac tcatactctt cctttttcaa tattattgaa gcatttatca gggttattgt
      661 ctcatgagcg gatacatatt tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc
      721 acatttcccc gaaaagtgcc acctgacgtc gacggatcgg gagatctccc gatcccctat
      781 ggtgcactct cagtacaatc tgctctgatg ccgcatagtt aagccagtat ctgctccctg
      841 cttgtgtgtt ggaggtcgct gagtagtgcg cgagcaaaat ttaagctaca acaaggcaag
      901 gcttgaccga caattgcatg aagaatctgc ttagggttag gcgttttgcg ctgcttcgcg
      961 atgtacgggc cagatatacg cgttgacatt gattattgac tagttattaa tagtaatcaa
     1021 ttacggggtc attagttcat agcccatata tggagttccg cgttacataa cttacggtaa
     1081 atggcccgcc tggctgaccg cccaacgacc cccgcccatt gacgtcaata atgacgtatg
     1141 ttcccatagt aacgccaata gggactttcc attgacgtca atgggtggag tatttacggt
     1201 aaactgccca cttggcagta catcaagtgt atcatatgcc aagtacgccc cctattgacg
     1261 tcaatgacgg taaatggccc gcctggcatt atgcccagta catgacctta tgggactttc
     1321 ctacttggca gtacatctac gtattagtca tcgctattac catggtgatg cggttttggc
     1381 agtacatcaa tgggcgtgga tagcggtttg actcacgggg atttccaagt ctccacccca
     1441 ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg ggactttcca aaatgtcgta
     1501 acaactccgc cccattgacg caaatgggcg gtaggcgtgt acggtgggag gtctatataa
     1561 gcagagctct ctggctaact agagaaccca ctgcttactg gcttatcgaa attaatacga
     1621 ctcactatag ggagacccaa gctagcatgg cctcctccga ggacgtcatc aaggagttca
     1681 tgcgcttcaa ggtgcgcatg gagggctccg tgaacggcca cgagttcgag atcgagggcg
     1741 agggcgaggg ccgcccctac gagggcaccc agaccgccaa gctgaaggtg accaagggcg
     1801 gccccctgcc cttcgcctgg gacatcctgt cccctcagtt ccagtacggc tccaaggcct
     1861 acgtgaagca ccccgccgac atccccgact acttgaagct gtccttcccc gagggcttca
     1921 agtgggagcg cgtgatgaac ttcgaggacg gcggcgtggt gaccgtgacc caggactcct
     1981 ccctgcagga cggcgagttc atctacaagg tgaagctgcg cggcaccaac ttcccctccg
     2041 acggccccgt aatgcagaag aagaccatgg gctgggaggc ctccaccgag cggatgtacc
     2101 ccgaggacgg cgccctgaag ggcgagatca agatgaggct gaagctgaag gacggcggcc
     2161 actacgacgc cgaggtcaag accacctaca tggccaagaa gcccgtgcag ctgcccggcg
     2221 cctacaagac cgacatcaag ctggacatca cctcccacaa cgaggactac accatcgtgg
     2281 aacagtacga gcgcgccgag ggccgccact ccaccggcgc ctccggactc agatctcgag
     2341 gcatggtgag caagggcgag gagctgttca ccggggtggt gcccatcctg gtcgagctgg
     2401 acggcgacgt aaacggccac aagttcagcg tgtccggcga gggcgagggc gatgccacct
     2461 acggcaagct gaccctgaag ttcatctgca ccaccggcaa gctgcccgtg ccctggccca
     2521 ccctcgtgac caccttcagc tacggcgtgc agtgcttcag ccgctacccc gaccacatga
     2581 agcagcacga cttcttcaag tccgccatgc ccgaaggcta cgtccaggag cgcaccatct
     2641 tcttcaagga cgacggcaac tacaagaccc gcgccgaggt gaagttcgag ggcgacacca
     2701 tggtgaaccg catcgagctg aagggcatcg acttcaagga ggacggcaac atcctggggc
     2761 acaagctgga gtacaagtac aacagccaca acgtctatat catggccgac aagcagaaga
     2821 acggcatcaa ggccaacttc aagatccgcc acaacatcga ggacggcagc gtgcagctcg
     2881 ccgaccacta ccagcagaac acccccatcg gcgacggccc cgtgctgctg cccgacaacc
     2941 actacctgag ccaccagtcc gccctgagca aagaccccaa cgagaagcgc gatcacatgg
     3001 tcctgctgga gttcgtgacc gccgccggga tcactctcgg catggacgag ctgtacaagg
     3061 aattcatggc tgccatccgg aagaaactgg tgattgttgg tgatggagcc tgtggaaaga
     3121 catgcttgct catagtcttc agcaaggacc agttcccaga ggtgtatgtg cccacagtgt
     3181 ttgagaacta tgtggcagat atcgaggtgg atggaaagca ggtagagttg gctttgtggg
     3241 acacagctgg gctggaagat tatgatcgcc tgaggcccct ctcctaccca gataccgatg
     3301 ttatactgat gtgtttttcc atcgacagcc ctgatagttt agaaaacatc ccagaaaagt
     3361 ggaccccaga agtcaagcat ttctgtccca acgtgcccat catcctggtt gggaataaga
     3421 aggatcttcg gaatgatgag cacacaaggc gggagctagc caagatgaag caggagccgg
     3481 tgaaacctga agaaggcaga gatatggcaa acaggattgg cgcttttggg tacatggagt
     3541 gttcagcaaa gaccaaagat ggagtgagag aggtttttga aatggctacg agagctgctc
     3601 tgcaagctag acgtgggaag aaaaaatctg gttgccttgt cttgtgactc gagcatgcat
     3661 ctagagggcc ctattctata gtgtcaccta aatgctagag ctcgctgatc agcctcgact
     3721 gtgccttcta gttgccagcc atctgttgtt tgcccctccc ccgtgccttc cttgaccctg
     3781 gaaggtgcca ctcccactgt cctttcctaa taaaatgagg aaattgcatc gcattgtctg
     3841 agtaggtgtc attctattct ggggggtggg gtggggcagg acagcaaggg ggaggattgg
     3901 gaagacaata gcaggcatgc tggggatgcg gtgggctcta tggcttctga ggcggaaaga
     3961 accagctggg gctctagggg gtatccccac gcgccctgta gcggcgcatt aagcgcggcg
     4021 ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca gcgccctagc gcccgctcct
     4081 ttcgctttct tcccttcctt tctcgccacg ttcgccggct ttccccgtca agctctaaat
     4141 cgggggctcc ctttagggtt ccgatttagt gctttacggc acctcgaccc caaaaaactt
     4201 gattagggtg atggttcacg tagtgggcca tcgccctgat agacggtttt tcgccctttg
     4261 acgttggagt ccacgttctt taatagtgga ctcttgttcc aaactggaac aacactcaac
     4321 cctatctcgg tctattcttt tgatttataa gggattttgc cgatttcggc ctattggtta
     4381 aaaaatgagc tgatttaaca aaaatttaac gcgaattaat tctgtggaat gtgtgtcagt
     4441 tagggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc atgcatctca
     4501 attagtcagc aaccaggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa
     4561 gcatgcatct caattagtca gcaaccatag tcccgcccct aactccgccc atcccgcccc
     4621 taactccgcc cagttccgcc cattctccgc cccatggctg actaattttt tttatttatg
     4681 cagaggccga ggccgcctct gcctctgagc tattccagaa gtagtgagga ggcttttttg
     4741 gaggcctagg cttttgcaaa aagctcccgg gagcttgtat atccattttc ggatctgatc
     4801 aagagacagg atgaggatcg tttcgcatga ttgaacaaga tggattgcac gcaggttctc
     4861 cggccgcttg ggtggagagg ctattcggct atgactgggc acaacagaca atcggctgct
     4921 ctgatgccgc cgtgttccgg ctgtcagcgc aggggcgccc ggttcttttt gtcaagaccg
     4981 acctgtccgg tgccctgaat gaactgcagg acgaggcagc gcggctatcg tggctggcca
     5041 cgacgggcgt tccttgcgca gctgtgctcg acgttgtcac tgaagcggga agggactggc
     5101 tgctattggg cgaagtgccg gggcaggatc tcctgtcatc tcaccttgct cctgccgaga
     5161 aagtatccat catggctgat gcaatgcggc ggctgcatac gcttgatccg gctacctgcc
     5221 cattcgacca ccaagcgaaa catcgcatcg agcgagcacg tactcggatg gaagccggtc
     5281 ttgtcgatca ggatgatctg gacgaagagc atcaggggct cgcgccagcc gaactgttcg
     5341 ccaggctcaa ggcgcgcatg cccgacggcg aggatctcgt cgtgacccat ggcgatgcct
     5401 gcttgccgaa tatcatggtg gaaaatggcc gcttttctgg attcatcgac tgtggccggc
     5461 tgggtgtggc ggaccgctat caggacatag cgttggctac ccgtgatatt gctgaagagc
     5521 ttggcggcga atgggctgac cgcttcctcg tgctttacgg tatcgccgct cccgattcgc
     5581 agcgcatcgc cttctatcgc cttcttgacg agttcttctg agcgggactc tggggttcga
     5641 aatgaccgac caagcgacgc ccaacctgcc atcacgagat ttcgattcca ccgccgcctt
     5701 ctatgaaagg ttgggcttcg gaatcgtttt ccgggacgcc ggctggatga tcctccagcg
     5761 cggggatctc atgctggagt tcttcgccca ccccaacttg tttattgcag cttataatgg
     5821 ttacaaataa agcaatagca tcacaaattt cacaaataaa gcattttttt cactgcattc
     5881 tagttgtggt ttgtccaaac tcatcaatgt atcttatcat gtctgtatac cgtcgacctc
     5941 tagctagagc ttggcgtaat catggtcata gctgtttcct gtgtgaaatt gttatccgct
     6001 cacaattcca cacaacatac gagccggaag cataaagtgt aaagcctggg gtgcctaatg
     6061 agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc gctttccagt cgggaaacct
     6121 gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg
     6181 gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc
     6241 ggtatcagct cactcaaagg cggtaatacg gttatccaca gaatcagggg ataacgcagg
     6301 aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct
     6361 ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac gctcaagtca
     6421 gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg gaagctccct
     6481 cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct ttctcccttc
     6541 gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt
     6601 tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct gcgccttatc
     6661 cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac tggcagcagc
     6721 cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt tcttgaagtg
     6781 gtggcctaac tacggctaca ctagaagaac agtatttggt atctgcgctc tgctgaagcc
     6841 agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca ccgctggtag
     6901 cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga
     6961 tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac gttaagggat
     7021 tttggtcatg agattatcaa aaaggatctt cacctagatc cttttaaatt aaaaatgaag
     7081 ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc aatgcttaat
     7141 cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg cctgactccc
     7201 cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagtg ctgcaatgat
     7261 accgcgagac ccacgctcac cggctccaga tttatcagca ataaaccagc cagccggaag
     7321 ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta tt
//
